Osteoclastogenesis from embryonic stem cells

胚胎干细胞的破骨细胞生成

• 批准号: 10670303
• 负责人: HAYASHI Shin-ichi
• 金额: $ 2.05万
• 依托单位: Tottori University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10670303/
• 关键词: osteoclast; embryonic stem cell; hematopoiesis; M-CSF; ODF; OPGL; endothelial cell; stromal cell; macrophage; ES細胞; tal-1; GATA-2

Osteoclasts are hematopoietic cells essential for bone resorption. To understand whole process of osteoclastogenesis, I have developed a culture system with a stromal cell line, in which promotes differentiation of hematopoietic cell lineage, and terminal differentiation of osteoclasts from single embryonic stem (ES) cells.This culture dispended with any passage or manipulation enabled us to investigate temporal and spatial localization of osteoclast lineage in the colonies from undifferentiated ES cells. On 4th day of cultures, hematopoietic cell and endothelial cell lineages arose in the central site of colonies. Cells expressed tartrate-resistant acid phosphatase (TRAPィイD1+ィエD1) which is a specific marker of osteoclast lineage were first detected on day 8 and subsequently localize at the edge of colonies and maturate into TRAPィイD1+ィエD1 multinucleated cells which have a potential of bone resorption.A potential of osteoclastogenesis of gene targeted ES cell lines in tal-1, Gata-1, Gata-2 and Fog genes was examined. Gata-1(-/-) and Fog(-/-) ES cells differentiate into osteoclasts normally. No osteoclasts were detected in the culture of tal-1(-/-) ES cells. Gata-2(-/-) ES cells developed mature osteoclasts but the frequency reduced to one-tenth of normal ES cell lines. I also decided the ordered requirement for osteoclastogenesis as tal-1 --> Gata --> macrophage colony-stimulating factor --> osteoclast differentiation factor.Recently, I established a new culture system for melanocyte development from ES cells. These systems are useful tools for study of osteoclastogenesis, because melanocytes are good internal control of cell dicision and culture conditions.

破骨细胞是骨吸收所必需的造血细胞。为了了解破骨细胞发生的全过程，我建立了一个由单个胚胎干细胞（ES细胞）诱导的破骨细胞分化为造血细胞系和终末分化的基质细胞系培养体系，这种培养不需要传代或操作，使我们能够研究破骨细胞系在未分化ES细胞集落中的时空定位。培养第4天，集落中心部位出现造血细胞和内皮细胞。在培养的第8天检测到破骨细胞系特异性标志物抗酒石酸酸性磷酸酶（TRAP β D1+ β D1）的表达，随后定位于集落边缘，成熟为具有骨吸收潜能的TRAP β D1+ β D1多核细胞，检测tal-1、加塔-1、加塔-2和Fog基因靶向ES细胞系的破骨细胞生成潜能。加塔-1（-/-）和Fog（-/-）ES细胞正常分化为破骨细胞。在tal-1（-/-）ES细胞培养中未检测到破骨细胞。加塔-2（-/-）ES细胞分化为成熟的破骨细胞，但分化频率降低至正常ES细胞系的十分之一。我还确定了破骨细胞生成的顺序为tal-1 ->加塔->巨噬细胞集落刺激因子->破骨细胞分化因子。这些系统是研究破骨细胞生成的有用工具，因为黑素细胞是细胞决策和培养条件的良好内部控制。

项目成果

S. Umeda et al.: "Deficiency of SHP-1 protein-tryosine phosphatase activity results in heightened osteoclast function and decreased bone density"Am. J. Pathol.. 155. 223-233 (1999)

S. Umeda 等人：“SHP-1 蛋白-酪氨酸磷酸酶活性的缺乏会导致破骨细胞功能增强和骨密度降低”Am。

T.Yamane et al.: "Sequential requirements of SCL/tal-1,GATA-2,M-CSF and osteoclast differentiation factor/osteoprotegerin ligand in osteoclast development"Exp.Hematol.. 印刷中.

T. Yamane 等人：“破骨细胞发育中 SCL/tal-1、GATA-2、M-CSF 和破骨细胞分化因子/骨保护素配体的顺序要求”Exp.Hematol.. 正在出版。

H.Yamazaki,et al.: "Promotion of early osteoclastogenesis and B lymphopoiesis in the bone marrow of transgenic rats with the env-pX gene of human T-cell lymphotropic virus type I." Oncogene. 17・23. 2955-2960 (1998)

H. Yamazaki 等人：“具有人类 T 细胞嗜淋巴细胞病毒 I 型 env-pX 基因的转基因大鼠的早期破骨细胞生成和 B 淋巴细胞生成的促进作用”（2955-2960）。 1998）

H. Tagaya et al.: "Intra- and extra-medullary B lymphopoiesis in osteopetrotic mice"Blood. (印刷中).

H. Tagaya 等人：“骨硬化小鼠的髓内和髓外 B 淋巴细胞生成”血液（正在出版）。

Tanio,Y., H.Yamazaki, T.Kunisada, K.Miyake, and S.I.Hayashi: "CD9 molecule expressed on stromal cells is involved in osteoclastogenesis"Exp. Hematol.. 27. 853-859 (1999)

Tanio,Y.、H.Yamazaki、T.Kunisada、K.Miyake 和 S.I.Hayashi：“基质细胞上表达的 CD9 分子参与破骨细胞生成”Exp。